Antioxidant and antiapoptotic properties of chlorogenic acid on human umbilical vein endothelial cells

Abstract

Chlorogenic acid (CGA) is one of the phenols isolated from Lonicera japonica Thumb. This study was designed to investigate whether CGA prevents human umbilical vein endothelial cells (HUVECs) from hydrogen peroxide (H2O2)–induced damage. The cell viability was examined by methylthiazolyl tetrazolium (MTT) assay. Malondialdehyde (MDA) level was measured using the thiobarbituric acid method. Reactive oxygen species (ROS) levels were determined using the 2',7'-dichlorofluorescin-diacetate (DCFH-DA) assay. The antioxidant enzymes activities, including glutathione peroxidase (GPx) and superoxide dismutase (SOD) were examined photometrically. Total antioxidant capacity (T-AOC) was measured using the ferric reducing activity of plasma (FRAP) assay. Apoptotic cells were detected by terminal deoxyribonucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL), annexin-V binding and by assessment of caspase-3. Protein and mRNA expression of Bax and Bcl-2 were determined by western blot and reverse transcription-polymerase chain reaction (RT-PCR). In the process of H2O2-induced damage, the expression of Bcl-2 was decreased and the expression of Bax was increased. The antioxidant enzymes activities and T-AOC levels were decreased. The content of MDA and the level of ROS were increased. Pretreatment with CGA significantly reduced oxidative stress by increased T-AOC levels and antioxidant enzymes activities, decreased ROS levels and lipid peroxidation. CGA significantly reduced apoptosis by up-regulation the expression of Bcl-2 and down-regulation the expression of Bax. Taken together, these results suggest that CGA exhibits antioxidant and antiapoptotic properties to prevent H2O2-induced cytotoxicity in HUVECs. Key words: Chlorogenic acid, human umbilical vein endothelial cells (HUVECs), hydrogen peroxide, oxidative stress, apoptosis.